Succinate Dehydrogenase B Subunit-Negative Jugular Foramen Paraganglioma Manifesting Malignant Progression with Pseudohypoxia-Related Atypical Uptake of [18F]-Fluoro-2-Deoxy-d-Glucose: A Case Report.

BACKGROUND: Paragangliomas are generally benign, slow-growing tumors. However, approximately 10%-20% are malignant, characterized by distant metastasis. Recently, a germ line mutation in succinate dehydrogenase B subunit (SDHB) has been shown to be associated with malignant behavior in paraganglioma. Here we present a case of SDHB-negative malignant paraganglioma of the jugular foramen with a pseudohypoxic microenvironment and unique imaging features on [18F]-fluoro-2-deoxy-d-glucose positron emission tomography ([18F]-FDG PET), and discuss the significance of SDHB immunohistochemistry and the potential of [18F]-FDG PET for clinical management. CASE DESCRIPTION: A 55-year-old woman was diagnosed with jugular foramen paraganglioma. Initial surgical resection was performed; however, follow-up [18F]-FDG PET indicated multiple uptake regions throughout the body. Biopsies for multiple recurrent lesions revealed consistent pathological features, suggesting distant metastasis. Immunohistochemical analysis revealed a lack of SDHB immunostaining in all specimens. Pseudohypoxic markers, including hypoxia-inducible factor-1α and downstream glycolysis enzymes, were strongly expressed. [18F]-FDG PET demonstrated increased uptake in the lesions, and the patient died 3 years after initial metastasis. CONCLUSION: In patients with head and neck paraganglioma without SDHB expression, close follow-up should be considered because of the risk for metastasis. In such cases, [18F]-FDG PET might be useful for detecting metastasis due to atypical accumulation from pseudohypoxia-induced glycolysis.

Renal cell carcinoma with angioleiomyoma-like stroma and clear cell papillary renal cell carcinoma: exploring SDHB protein immunohistochemistry and the relationship to tuberous sclerosis complex.

Renal cell carcinoma (RCC) with angioleiomyoma-like stroma appears to be molecularly distinct from clear cell RCC; however, its relationship to clear cell papillary RCC remains debated. Recent studies have found that similar tumors sometimes occur in patients with tuberous sclerosis complex (TSC), of which 1 study found unexpectedly negative succinate dehydrogenase B (SDHB) immunostaining. We evaluated immunohistochemistry for SDHB in 12 apparently sporadic RCCs with angioleiomyoma-like stroma and correlated with clinical information for stigmata of TSC. Tumors were compared with a group of 16 clear cell papillary RCCs and 6 unclassified tumors with prominent stroma. With the exception of 1 unclassified tumor, all exhibited at least focal cytoplasmic staining for SDHB protein, often requiring high magnification and better appreciated with increased antibody concentration. Detailed history information was available for 9 of 11 patients with smooth muscle-rich tumors, revealing no stigmata of undiagnosed TSC. Electron microscopy performed on 1 of these tumors revealed mitochondria to be very sparse, potentially accounting for the weak immunohistochemical labeling for SDHB protein. Weak SDHB immunostaining may represent another shared feature of RCC with angioleiomyoma-like stroma and clear cell papillary RCC, likely due to sparse mitochondria, strengthening the possible relationship of these entities. Although smooth muscle-rich tumors have been recently reported in patients with TSC, absence of staining in tumors with this pattern may not be specific for TSC. In tumors with pale or clear cytoplasm, immunohistochemical staining for SDHB should be interpreted with caution as evidence of abnormality in the SDH pathway.

The utility of SDHB and FH immunohistochemistry in patients evaluated for hereditary paraganglioma-pheochromocytoma syndromes.

A significant portion of paragangliomas (PGL) and pheochromocytomas (PCC) occur in patients with hereditary PGL/PCC syndromes, including those with germline mutations in succinate dehydrogenase (SDHx) subunit genes. Recently, germline fumarate hydratase (FH) mutations have been identified in a subset of PGL/PCC, and patients with hereditary leiomyomatosis and renal cell carcinoma (HLRCC) may have an increased risk of developing PGL/PCC. SDHB immunohistochemistry (IHC) has previously been shown to be useful for identifying SDHx-deficient PGL/PCC, however, FH IHC has never been explored in these tumors. Thus, we characterized SDHB and FH IHC in a large cohort of PGL/PCC patients (n = 41) at our institution who were evaluated for hereditary PGL/PCC syndromes. Overall, there was strong, positive correlation between germline SDHx subunit gene mutation status and SDHB IHC status (rφ = 0.77; P < .0001), with high corresponding sensitivity, specificity, positive predictive value, and negative predictive value (95.0%, 81.8%, 82.6%, and 94.7%, respectively). Although SDHB loss by IHC was highly correlated with germline SDHx gene mutations, its utility in this population was dependent on clinicopathologic context: while all head and neck PGL patients with SDHB-deficient tumors had germline SDHx gene mutations, only a small subset (25.0%) of PCC patients with SDHB-deficient tumors harbored a germline SDHx gene mutation. Finally, although our cohort contained only one HLRCC patient, their tumor was FH-deficient by IHC, and all other PGL/PCC showed retained FH IHC. Thus, in the appropriate clinical setting, SDHB and FH IHC may be useful for identifying PGL/PCC patients for Medical Genetics evaluation.

Composite Pheochromocytoma/Paraganglioma-Ganglioneuroma: A Clinicopathologic Study of Eight Cases with Analysis of Succinate Dehydrogenase.

Ganglioneuromas represent the most well-differentiated spectrum of neoplasia arising from the sympathetic nervous system, while neuroblastomas represent the most poorly differentiated counterpart, and ganglioneuroblastomas represent intermediate stages of differentiation. Small series of cases have documented the co-occurrence of ganglioneuroma with a pheochromocytoma (Pheo)/paraganglioma (PGL) component. We report the clinicopathologic features of eight such cases, diagnosed between 2003 and 2015 with a mean follow-up of 22 months (1-47), which were evaluated for syndrome associations, SDHB expression, and clinical outcome. Mutations of the succinate dehydrogenase (SDH) complex subunits (A, B, C, D, and SDHAF2) have been implicated in predicting metastatic behavior and in identifying possible paraganglioma syndromes. The proliferative index was calculated by manual quantification of Ki-67-positive cells at selected hot-spots using ImageJ (NIH). In our series, composite Pheo/PGL-ganglioneuromas predominantly involved the adrenal gland (Pheo 7, PGL 1). The cases had an equal gender distribution (males 4, females 4), with a mean age at diagnosis of 67 years (range 53 to 86 years), an average size of 5.2 cm (range 2 to 8.2 cm), an average weight of 49.3 g (7.8 to 144.7 g, n = 6), and the majority were functionally active (7 of 8, 88%). The mean Ki67 proliferation rate was 2% (range 0.3 to 3%), and all cases retained SDHB expression (8/8, 100%). No patient (0/8, 0%) developed metastatic disease on follow-up. One patient had a retroperitoneal composite PGL-ganglioneuroma in the setting of neurofibromatosis type 1. No recurrent disease or other associations were identified. In our study, composite Pheo/PGL-ganglioneuromas predominantly affected the adrenal gland in older patients, showed no loss of SDHB, and no disease recurrence was identified.

Biogenesis and functions of mammalian iron-sulfur proteins in the regulation of iron homeostasis and pivotal metabolic pathways.

Fe-S cofactors are composed of iron and inorganic sulfur in various stoichiometries. A complex assembly pathway conducts their initial synthesis and subsequent binding to recipient proteins. In this minireview, we discuss how discovery of the role of the mammalian cytosolic aconitase, known as iron regulatory protein 1 (IRP1), led to the characterization of the function of its Fe-S cluster in sensing and regulating cellular iron homeostasis. Moreover, we present an overview of recent studies that have provided insights into the mechanism of Fe-S cluster transfer to recipient Fe-S proteins.

Cancer-associated fibroblasts enhance pancreatic cancer cell invasion by remodeling the metabolic conversion mechanism.

We investigated the mechanism of cancer-associated fibroblasts (CAFs) in promoting the invasion and metastasis of pancreatic cancer cells in a non-vascular manner. We verified the original generation of isolated cultured CAFs and normal fibroblasts (NFs) based on the expression of α-SMA and vimentin, and we examined the cell glycolysis level through glucose consumption and lactate production experiments. The mRNA and protein expression of CAF glycolytic enzymes, lactate dehydrogenase and pyruvate kinase m2, were examined by RT-PCR and western blotting, respectively. In vitro culture first-generation pancreatic CAFs were collected and cultured together with pancreas cancer BxPc-3 and Panc-1 cells. Cell invasion and migration were assessed using a Transwell assay and scratch test, respectively. Mitochondrial activity was assessed by experimentally determining oxidative phosphorylation (OP) activity. The aerobic oxidation index of cancer cells was also examined. Succinate dehydrogenase, fumarate hydratase (FH), and monocarboxylate transporter 1 (MCT1) expression were examined using an MCT1-specific inhibitor to remove 'tumor-stromal' metabolic coupling to observe the influence of cell interstices on pancreas cancer progression. First-generation isolated cultured CAFs and NFs both grew well, and showed active proliferation. Glucose absorption and lactate production were significantly enhanced in CAFs compared with that in NFs. PCR and western blotting showed that the lactate dehydrogenase and pyruvate kinase m2 mRNA and protein expression levels were increased in the CAFs. After indirect co-culture, OP was increased in the BxPc-3 and Panc-1 cells; correspondingly, succinate dehydrogenase, FH and MCT expression were increased. After the MCT1-specific inhibitor removed 'tumor-stromal' metabolic coupling, the migration and invasion abilities of the pancreatic cancer cells were decreased. Pancreatic CAFs can alter metabolism as well as communicate with and respond to cancer cell migration and invasion. This may be an important mechanism for promoting tumor progression in a non-vascular manner in the tumor microenvironment. The mechanism by which CAFs reshape metabolic transition requires further analysis.

Potential Pitfalls of SDH Immunohistochemical Detection in Paragangliomas and Phaeochromocytomas Harbouring Germline SDHx Gene Mutation.

BACKGROUND/AIM: Germline mutations in any of the succinate dehydrogenase (SDH) genes result in destabilization of the SDH protein complex and loss of SDHB expression at immunohistochemistry. SDHA is lost together with SDHB in SDHA-mutated tumours, but its expression is retained in tumours with other SDH mutations. We investigated whether SDHA/SDHB immunohistochemistry is able to identify SDH-related tumours in a retrospective case series of phaeochromocytomas (PCCs) and paragangliomas (PGLs). MATERIALS AND METHODS: SDHA and SDHB immunostaining was performed in 13 SDH gene-mutated tumours (SDHB: n=3; SDHC: n=1; SDHD: n=9) and 16 wild-type tumours. Protein expression by western blot analysis and enzymatic activity were also assessed. RESULTS: Tumours harbouring SDH gene mutations demonstrated a significant reduction in enzymatic activity and protein expression when compared to wild-type tumours. SDHB immunostaining detected 76.9% of SDH mutated PCCs/PGLs (3/3 SDHB-mutated samples; 1/1 SDHC-mutated sample; 6/9 SDHD-mutated samples). In three SDHD-related tumours with the same mutation (p.Pro81Leu), positive (n=2) or weakly diffuse (n=1) SDHB staining was observed. All wild-type PCCs/PGLs exhibited SDHB immunoreactivity, while immunostaining for SDHA was positive in 93.8% cases and weakly diffuse in one (6.2%). SDHA protein expression was preserved in all tumours with mutations. CONCLUSION: SDHA and SDHB immunohistochemistry should be interpreted with caution, due to possible false-positive or false-negative results, and ideally in the setting of quality assurance provided by molecular testing. In SDHD mutation, weak non-specific cytoplasmic staining occurs commonly, and this pattern of staining can be difficult to interpret with certainty.

Use of antisense oligonucleotides to correct the splicing error in ISCU myopathy patient cell lines.

ISCU myopathy is an inherited disease that primarily affects individuals of northern Swedish descent who share a single point mutation in the fourth intron of the ISCU gene. The current study shows correction of specific phenotypes associated with disease following treatment with an antisense oligonucleotide (ASO) targeted to the site of the mutation. We have shown that ASO treatment diminished aberrant splicing and increased ISCU protein levels in both patient fibroblasts and patient myotubes in a concentration dependent fashion. Upon ASO treatment, levels of SDHB in patient myotubular cell lines increased to levels observed in control myotubular cell lines. Additionally, we have shown that both patient fibroblast and myotubular cell lines displayed an increase in complex II activity with a concomitant decrease in succinate levels in patient myotubular cell lines after ASO treatment. Mitochondrial and cytosolic aconitase activities increased significantly following ASO treatment in patient myotubes. The current study suggests that ASO treatment may serve as a viable approach to correcting ISCU myopathy in patients.

Urinary Bladder Paragangliomas: Analysis of Succinate Dehydrogenase and Outcome.

Paragangliomas of the urinary bladder can arise sporadically or as a part of hereditary syndromes including those with underlying mutations in the succinate dehydrogenase (SDH) genes, which serve as tumor suppressors. SDH deficiency can be screened for by absence of immunohistochemical detection of SDHB. In this study of 11 cases, clinical follow-up was available for 9/11 cases. The cases were reviewed and graded based on the grading system for adrenal pheochromocytomas and paragangliomas (GAPP) criteria. Immunohistochemistry was performed for Ki67 and SDHB. Proliferative index was calculated by quantification of Ki67-positive cells at hot spots. The medical record was accessed for documentation of germline SDH mutations. Urinary bladder paragangliomas had a female predilection (8/11 cases), and 5/11 cases exhibited metastatic behavior. Patients with metastatic disease tended to be younger (mean age 43 vs 49 years), have larger lesions (5.8 vs 1.5 cm), and presented with catecholamine excess (4/4 vs 2/6 patients with non-metastatic lesions). Patients with metastatic disease had a higher mean Ki67 proliferation rate (4.9 vs 1.3 %) and GAPP score (mean of 5.8 vs 3.8) (p = 0.01). IHC for SDHB expression revealed loss of expression in 2/6 cases of non-metastatic paragangliomas compared to 4/5 patients with metastatic paragangliomas. Interestingly, of these four patients, two had a documented mutation of SDHB, one patient had a SDHC mutation, and another patient had a history of familial disease without mutation analysis being performed. Our study, suggests that SDH loss was suggestive of metastatic behavior in addition to younger age at diagnosis, larger tumor size, and higher Ki67 proliferation rate and catecholamine type.

Down-regulation of succinate dehydrogenase subunit B and up-regulation of pyruvate dehydrogenase kinase 1 predicts poor prognosis in recurrent nasopharyngeal carcinoma.

Succinate dehydrogenase subunit B (SDHB) and pyruvate dehydrogenase kinase 1 (PDK1) play key roles in the regulation of growth and survival of various cancers. This study aimed to investigate expression of SDHB and PDK1 in recurrent nasopharyngeal carcinoma (rNPC) tissues and analyzed the association of SDHB and PDK1 expression with the clinical significance and potential prognostic implication of rNPC. Immunohistochemistry was performed to determine the expression of SDHB and PDK1 in tissues in primary NPC (pNPC) and rNPC patients. Our results revealed that expression of SDHB in rNPC was significantly lower than that in pNPC, while the expression of PDK1 was higher compared to pNPC. The expression levels of SDHB and PDK1 were associated with T stage, N stage, clinical stage, and metastasis of rNPC. Survival analysis showed that patients with low SDHB expression had a significantly shorter overall survival time than those with high SDHB expression. Patients with high PDK1 expression had a shorter survival time than patients with low PDK1 expression. Multivariate analysis showed that the expression of SDHB and PDK1 was an independent predictor for the survival of patients with rNPC. Our results demonstrated that down-regulation of SDHB and up-regulation of PDK1 may be novel biomarkers for predicting advanced tumor progression and unfavorable prognosis in rNPC patients.

Succinate dehydrogenase (SDH)-deficient renal carcinoma: a morphologically distinct entity: a clinicopathologic series of 36 tumors from 27 patients.

Succinate dehydrogenase (SDH)-deficient renal carcinoma has been accepted as a provisional entity in the 2013 International Society of Urological Pathology Vancouver Classification. To further define its morphologic and clinical features, we studied a multi-institutional cohort of 36 SDH-deficient renal carcinomas from 27 patients, including 21 previously unreported cases. We estimate that 0.05% to 0.2% of all renal carcinomas are SDH deficient. Mean patient age at presentation was 37 years (range, 14 to 76 y), with a slight male predominance (M:F=1.7:1). Bilateral tumors were observed in 26% of patients. Thirty-four (94%) tumors demonstrated the previously reported morphology at least focally, which included: solid or focally cystic growth, uniform cytology with eosinophilic flocculent cytoplasm, intracytoplasmic vacuolations and inclusions, and round to oval low-grade nuclei. All 17 patients who underwent genetic testing for mutation in the SDH subunits demonstrated germline mutations (16 in SDHB and 1 in SDHC). Nine of 27 (33%) patients developed metastatic disease, 2 of them after prolonged follow-up (5.5 and 30 y). Seven of 10 patients (70%) with high-grade nuclei metastasized as did all 4 patients with coagulative necrosis. Two of 17 (12%) patients with low-grade nuclei metastasized, and both had unbiopsied contralateral tumors, which may have been the origin of the metastatic disease. In conclusion, SDH-deficient renal carcinoma is a rare and unique type of renal carcinoma, exhibiting stereotypical morphologic features in the great majority of cases and showing a strong relationship with SDH germline mutation. Although this tumor may undergo dedifferentiation and metastasize, sometimes after a prolonged delay, metastatic disease is rare in the absence of high-grade nuclear atypia or coagulative necrosis.

The expression of succinate dehydrogenase in breast phyllodes tumor.

The purpose of this study is to investigate the expression of succinate dehydrogenase (SDH)A, SDHB, and HIF-1α in phyllodes tumors and the association with clinic-pathologic factors. Using tissue microarray (TMA) for 206 phyllodes tumor cases, we performed immunohistochemical stains for SDHA, SDHB, and HIF-1α and analyzed their expression in regard to clinicopathologic parameters of each case. The cases were comprised of 156 benign, 34 borderline, and 16 malignant phyllodes tumors. The expression of stromal SDHA and epithelial- and stromal- SDHB increased as the tumor progressed from benign to malignant (P⟨0.001). There were five stromal SDHA-negative cases and 31 stromal SDHB-negative cases. SDHB negativity was associated with a lower histologic grade (P=0.054) and lower stromal atypia (P=0.048). Univariate analysis revealed that a shorter disease free survival (DFS) was associated with stromal SDHB high-positivity (P=0.013) and a shorter overall survival (OS) was associated with high-positivity of stromal SDHA and SDHB (P⟨0.001 and P⟨0.001, respectively). The multivariate Cox analysis with the variables stromal cellularity, stromal atypia, stromal mitosis, stromal overgrowth, tumor margin, stromal SDHA expression, and stromal SDHB expression revealed that stromal overgrowth was associated with a shorter DFS (hazard ratio: 24.78, 95% CI: 3.126-196.5, P=0.002) and a shorter OS (hazard ratio: 176.7, 95% CI: 8.466-3691, P=0.001). In conclusion, Tumor grade is positively correlated with SDHA and SDHB expression in the tumor stroma in phyllodes tumors of the breast. This result may be attributed to the increased metabolic demand in high grade tumors.

Analysis of covalent flavinylation using thermostable succinate dehydrogenase from Thermus thermophilus and Sulfolobus tokodaii lacking SdhE homologs.

Recent studies have indicated that post-translational flavinylation of succinate dehydrogenase subunit A (SdhA) in eukaryotes and bacteria require the chaperone-like proteins Sdh5 and SdhE, respectively. How does covalent flavinylation occur in prokaryotes, which lack SdhE homologs? In this study, I showed that covalent flavinylation in two hyperthermophilic bacteria/archaea lacking SdhE, Thermus thermophilus and Sulfolobus tokodaii, requires heat and dicarboxylic acid. These thermophilic bacteria/archaea inhabit hot environments and are said to be genetically far removed from mesophilic bacteria which possess SdhE. Since mesophilic bacteria have been effective at covalent bonding in temperate environments, they may have caused the evolution of SdhE.

The succinate dehydrogenase assembly factor, SdhE, is required for the flavinylation and activation of fumarate reductase in bacteria.

The activity of the respiratory enzyme fumarate reductase (FRD) is dependent on the covalent attachment of the redox cofactor flavin adenine dinucleotide (FAD). We demonstrate that the FAD assembly factor SdhE, which flavinylates and activates the respiratory enzyme succinate dehydrogenase (SDH), is also required for the complete activation and flavinylation of FRD. SdhE interacted with, and flavinylated, the flavoprotein subunit FrdA, whilst mutations in a conserved RGxxE motif impaired the complete flavinylation and activation of FRD. These results are of widespread relevance because SDH and FRD play an important role in cellular energetics and are required for virulence in many important bacterial pathogens.

Analysis of all subunits, SDHA, SDHB, SDHC, SDHD, of the succinate dehydrogenase complex in KIT/PDGFRA wild-type GIST.

Mutations of genes encoding the subunits of the succinate dehydrogenase (SDH) complex were described in KIT/PDGFRA wild-type GIST separately in different reports. In this study, we simultaneously sequenced the genome of all subunits, SDHA, SDHB, SDHC, and SDHD in a larger series of KIT/PDGFRA wild-type GIST in order to evaluate the frequency of the mutations and explore their biological role. SDHA, SDHB, SDHC, and SDHD were sequenced on the available samples obtained from 34 KIT/PDGFRA wild-type GISTs. Of these, in 10 cases, both tumor and peripheral blood (PB) were available, in 19 cases only tumor, and in 5 cases only PB. Overall, 9 of the 34 patients with KIT/PDGFRA wild-type GIST carried mutations in one of the four subunits of the SDH complex (six patients in SDHA, two in SDHB, one in SDHC). WB and immunohistochemistry analysis showed that patients with KIT/PDGFRA wild-type GIST who harbored SDHA mutations exhibited a significant downregulation of both SDHA and SDHB protein expression, with respect to the other GIST lacking SDH mutations and to KIT/PDGFRA-mutated GIST. Clinically, four out of six patients with SDHA mutations presented with metastatic disease at diagnosis with a very slow, indolent course. Patients with KIT/PDGFRA wild-type GIST may harbor germline and/or de novo mutations of SDH complex with prevalence for mutations within SDHA, which is associated with a downregulation of SDHA and SDHB protein expression. The presence of germline mutations may suggest that these patients should be followed up for the risk of development of other cancers.

Comparative study of changes in energy metabolism in rat cardiomyocytes in postinfarction cardiosclerosis and diabetes mellitus.

Cardiomyocyte energy metabolism in experimental unfolding postinfarction cardiosclerosis and diabetes mellitus was studied. Postinfarction cardiosclerosis formed 6 weeks after coronary artery occlusion. Diabetes mellitus was induced by intraperitoneal injection of streptozotocin (60 mg/kg). The rate of oxygen consumption in postinfarction cardiosclerosis and diabetes increased by 3.4 and 4.2 times, respectively. Stimulation of mitochondrial respiration (ATP, palmitic acid) significantly increased oxygen consumption in animals with postinfarction cardiosclerosis and significantly reduced this process in diabetes. The content of LDH and SDH in the myocardium of animals with diabetes and postinfarction cardiosclerosis was significantly below the control. Hence, the development of postinfarction cardiosclerosis and diabetes mellitus were characterized by reduced generation of ATP in anaerobic and aerobic pathways and oxidative phosphorylation in cardiomyocytes.

Control of RNA stability by NrrF, an iron-regulated small RNA in Neisseria gonorrhoeae.

Regulation of gene expression by small noncoding RNAs (sRNAs) plays a critical role in bacterial response to physiological stresses. NrrF, a trans-acting sRNA in Neisseria meningitidis and Neisseria gonorrhoeae, has been shown in the meningococcus to control indirectly, in response to iron (Fe) availability, the transcription of genes encoding subunits of succinate dehydrogenase, a Fe-requiring enzyme. Given that in other organisms, sRNAs target multiple mRNAs to control gene expression, we used a global approach to examine the role of NrrF in controlling gonococcal transcription. Three strains, including N. gonorrhoeae FA1090, an nrrF deletion mutant, and a complemented derivative, were examined using a custom CombiMatrix microarray to assess the role of this sRNA in controlling gene expression in response to Fe availability. In the absence of NrrF, the mRNA half-lives for 12 genes under Fe-depleted growth conditions were longer than those in FA1090. The 12 genes controlled by NrrF encoded proteins with biological functions including energy metabolism, oxidative stress, antibiotic resistance, and amino acid synthesis, as well as hypothetical proteins and a regulatory protein whose functions are not fully understood.

Identification of succinate dehydrogenase-deficient bladder paragangliomas.

A significant number of patients with paragangliomas harbor germline mutations in one of the succinate dehydrogenase (SDH) genes (SDHA, B, C, or D). Tumors with mutations in SDH genes can be identified using immunohistochemistry. Loss of SDHB staining is seen in tumors with a mutation in any one of the SDH genes, whereas loss of both SDHB and SDHA expression is seen only in the context of an SDHA mutation. Identifying an SDH-deficient tumor can be prognostically significant, as tumors with SDHB mutations are more likely to pursue a malignant course. Although the rate of SDH deficiency in paragangliomas in general is known to be approximately 30%, there are only rare reports of SDH-deficient bladder paragangliomas. Therefore, the aim of this study was to determine the rate of SDH deficiency in bladder paragangliomas. Eleven cases of bladder paragangliomas were identified. Hematoxylin and eosin-stained slides of all tumors were reviewed, and immunohistochemical analysis for SDHB and SDHA was performed. For cases with loss of SDHA expression by immunohistochemistry, mutation analysis of the SDHA gene was performed. Loss of SDHB staining was seen in 3 (27%) cases (2 with loss of SDHB only, 1 with loss of SDHB and SDHA). Patients with SDH-deficient tumors were younger than those with tumors with intact SDH expression (mean age at presentation 39 y and 58 y, respectively). Of the 2 patients with SDHB-deficient and SDHA-intact tumors, one was found to have a germline SDHB mutation, and the other had a family history of a malignant paraganglioma. Both patients developed metastatic disease. The one patient with a tumor that was deficient for both SDHB and SDHA had no family history of paragangliomas and no evidence of metastatic disease. Sequencing of this tumor revealed a deleterious heterozygous single-base pair substitution in exon 10 of SDHA (c.1340 A>G; p.His447Arg) in both the tumor and normal tissue, indicative of a germline SDHA mutation, and a deleterious single-base pair substitution in exon 5 of SDHA (c.484 A>T; p.Arg162*) in 1 allele of the tumor only. No patients with intact SDH expression had a family history of paragangliomas; 1 had a synchronous paraganglioma, but none developed metastatic disease. A significant subset of bladder paragangliomas is SDH deficient. It is essential to identify SDH-deficient tumors, as the presence of an SDH mutation has prognostic implications and is important in guiding genetic counseling.

Effects of zoledronic acid on odontoblast-like cells.

The aim of the study was to evaluate the effects of a highly potent bisphosphonate, zoledronic acid (ZOL), on cultured odontoblast-like cells MDPC-23. The cells (1.5×10(4)cells/cm(2)) were seeded for 48h in wells of 24-well dished. Then, the plain culture medium (DMEM) was replaced by fresh medium without fetal bovine serum. After 24h, ZOL (1 or 5µM) was added to the medium and maintained in contact with the cells for 24h. After this period, the succinic dehydrogenase (SDH) enzyme production (cell viability - MTT assay), total protein (TP) production, alkaline phosphatase (ALP) activity, and gene expression (qPCR) of collagen type I (Col-I) and ALP were evaluated. Cell morphology was assessed by SEM. Five µM ZOL caused a significant decrease in SDH production. Both ZOL concentrations caused a dose-dependent significant decrease in TP production and ALP activity. ZOL also produced discret morphological alterations in the MDPC-23 cells. Regarding gene expression, 1µM ZOL caused a significant increase in Col-I expression. Although 5µM ZOL did not affect Col-I expression, it caused a significant alteration in ALP expression (ANOVA and Tukey's test, p<0.05). ZOL presented a dose-dependent cytotoxic effect on the odontoblast-like cells, suggesting that under clinical conditions the release of this drug from dentin could cause damage to the pulpo-dentin complex.

Angiotensin receptor blockade recovers hepatic UCP2 expression and aconitase and SDH activities and ameliorates hepatic oxidative damage in insulin resistant rats.

Metabolic syndrome (MetS) is commonly associated with elevated renin-angiotensin system, oxidative stress, and steatohepatitis with down-regulation of uncoupling proteins (UCPs). However, the mechanisms linking renin-angiotensin system, steatosis, and UCP2 to hepatic oxidative damage during insulin resistance are not described. To test the hypothesis that angiotensin receptor activation contributes to decreased hepatic UCP2 expression and aconitase activity and to increased oxidative damage after increased glucose intake in a model of MetS, lean and obese Long Evans rats (n = 10/group) were randomly assigned to the following groups: 1) untreated Long Evans Tokushima Otsuka (lean, strain control), 2) untreated Otsuka Long Evans Tokushima Fatty (OLETF) (MetS model), 3) OLETF + angiotensin receptor blocker (ARB) (10 mg olmesartan/kg·d × 6 wk), 4) OLETF + high glucose (HG) (5% in drinking water × 6 wk), and 5) OLETF + ARB + HG (ARB/HG × 6 wk). HG increased body mass (37%), plasma triglycerides (TGs) (35%), plasma glycerol (87%), plasma free fatty acids (28%), and hepatic nitrotyrosine (74%). ARB treatment in HG decreased body mass (12%), plasma TG (15%), plasma glycerol (23%), plasma free fatty acids (14%), and hepatic TG content (42%), suggesting that angiotensin receptor type 1 (AT1) activation and increased adiposity contribute to the development of obesity-related dyslipidemia. ARB in HG also decreased hepatic nitrotyrosine and increased hepatic UCP2 expression (59%) and aconitase activity (40%), as well as antioxidant enzyme activities (50-120%), suggesting that AT1 activation also contributes to protein oxidation, impaired lipid metabolism, and antioxidant metabolism in the liver. Thus, in addition to promoting obesity-related hypertension, AT1 activation may also impair lipid metabolism and antioxidant capacity, resulting in steatosis via decreased UCP2 and tricarboxylic acid cycle activity.